A comprehensive review of nondermatophyte mould onychomycosis: Epidemiology, diagnosis and management.

Nondermatophyte moulds (NDMs) are widely distributed and can be detected in association with mycotic nails; however, sometimes it can be challenging to establish the role of NDMs in the pathogenesis of onychomycosis (i.e. causative vs. contaminant). In studies where the ongoing invasive presence of NDMs is confirmed through repeat cultures, the global prevalence of NDMs in onychomycosis patients is estimated at 6.9% with the 3 most common genus being: Aspergillus, Scopulariopsis and Fusarium. NDM onychomycosis can, in many cases, appear clinically indistinguishable from dermatophyte onychomycosis. Clinical features suggestive of NDMs include proximal subungual onychomycosis with paronychia associated with Aspergillus spp., Fusarium spp. and Scopulariopsis brevicaulis, as well as superficial white onychomycosis in a deep and diffused pattern associated with Aspergillus and Fusarium. Longitudinal streaks seen in patients with distal and lateral onychomycosis may serve as an additional indicator. For diagnosis, light microscopic examination should demonstrate fungal filaments consistent with an NDM with at least two independent isolations in the absence of a dermatophyte; the advent of molecular testing combined with histological assessment may serve as an alternative with improved sensitivity and turnover time. In most instances, antifungal susceptibility testing has limited value. Information on effective treatments for NDM onychomycosis is relatively scarce, unlike the situation in the study of dermatophyte onychomycosis. Terbinafine and itraconazole therapy (continuous and pulsed) appear effective to varying extents for treating onychomycosis caused by Aspergillus, Fusarium or Scopulariopsis. There is scant literature on oral treatments for Neoscytalidium.

Terbinafine Resistant Trichophyton Indotineae Isolated in Patients With Superficial Dermatophyte Infection in Canadian Patients.

BACKGROUND: Trichophyton mentagrophytes type VIII (segregated as Trichophyton indotineae) is a new strain of antifungal resistant Trichophyton spp. that has been found in different countries around the world. This new strain has been found to be resistant to terbinafine. OBJECTIVES: We present the clinical characteristics, diagnosis and treatment approach of Canadian patients with mycology and molecular confirmation of superficial mycosis caused by T. indotineae. METHODS: Mycology testing from cultures and PCR were used to confirm T. indotineae. We collected clinical information from patients with a confirmed diagnosis. RESULTS: We report eight Canadian patients mainly presenting with extensive superficial mycosis due to T. indotineae. Three patients presented lesions on the face in addition to lesions on their body. Four patients were initially started on itraconazole or fluconazole, one patient was started on topical therapy, and three patients were referred to infectious disease. CONCLUSIONS: This new strain represents a dermatology and public health concern. Treatment guidelines are lacking. We include a practical approach and treatment recommendations for clinicians who will be encountering these emerging cases in Canada while waiting for evidence-based treatment guidelines.

Assessing the Use of DNA Detection Platforms Combined with Passive Wind-Powered Spore Traps for Early Surveillance of Potato and Tomato Late Blight in Canada.

Quantitative PCR (qPCR), loop-mediated amplification (LAMP), and lateral flow strip-based recombinase polymerase amplification (RPA-LFS) assays were assessed for early detection of Phytophthora infestans, the global causal agent of potato and tomato late blight, on passive wind-powered spore traps known as Spornados. Spore traps were deployed in potato and tomato fields during the 2018, 2019, and 2020 growing seasons in the provinces of Alberta, British Columbia, Manitoba, Prince Edward Island, and Ontario. All assays used DNA extracts from Spornado cassette membranes targeting the P. infestans nuclear ribosomal internal transcribed spacer. A total of 1,003 Spornado samples were qPCR tested, yielding 115 positive samples for P. infestans spores. In further assessment of these samples, LAMP detected P. infestans in 108 (93.9%) of 115 qPCR positive samples, and RPA-LFS detected it in 103 (89.6%). None of the assays showed cross-reaction with other Phytophthora species or pathogenic fungi known to infect potato and tomato. The qPCR detected ≤1 fg of P. infestans DNA, and LAMP and RPA-LFS amplified 10 fg in as little as 10 min. All assays detected P. infestans before the first report of late blight symptoms in commercial potato or tomato fields within each region or province. The combination of Spornado passive samplers with qPCR, LAMP, or RPA-LFS proved a valuable spore trapping system for early surveillance of late blight in potato and tomato. Both LAMP and RPA-LFS showed potential as alternative approaches to qPCR for in-field monitoring of P. infestans.

Evaluation of the Copan ESwab Transport System for Viability of Pathogenic Fungi by Use of a Modification of Clinical and Laboratory Standards Institute Document M40-A2.

The Copan ESwab system was evaluated for its ability to maintain the viability of pathogenic fungi. Tests followed the Clinical and Laboratory Standards Institute (CLSI) document M40-A2 roll-plate method at room and refrigerator temperatures. A system was devised for standardizing homogeneous inoculum suspensions of variously sized conidia and sporangiospores of filamentous fungi. A total of 19 clinical and reference strains were standardized to a 0.5 McFarland turbidity standard with a simple photometer. Corresponding optical densities were measured with a spectrophotometer. Colony counts equal to or greater than those seen at time zero were obtained for the entire test panel. Results indicate that the Copan ESwab system effectively maintains prevalent opportunistic fungal organisms for at least 48 h.

Phaeoacremonium tuscanicum and Phaeoacremonium indicum sp. nov. associated with subcutaneous phaeohyphomycosis.

Two cases of phaeohyphomycotic infections were caused by Phaeoacremonium tuscanicum, not previously identified in human infections, and one new species, Phaeoacremonium indicum, respectively. Morphological and cultural investigation as well as phylogenetic analysis was constructed based on maximum likelihood analyses using actin and -tubulin sequences to identify the fungal isolates.

Phialemoniopsis, a new genus of Sordariomycetes, and new species of Phialemonium and Lecythophora.

In molecular studies involving numerous clinical isolates of the genera Acremonium, Phialemonium and Lecytophora some of them could not be identified. To clarify the phylogenetic relationships among these fungi and other related taxa, we performed a polyphasic study based on a detailed morphological study and on the analysis of sequences of four loci: the internal transcribed spacer regions, the D1/D2 domains of the 28S rRNA, actin and ß-tubulin genes. The combination of the resulting data let us propose the new genus Phialemoniopsis to accommodate the opportunistic fungi Phialemonium curvatum and Sarcopodium oculorum and two new species, Phialemoniopsis cornearis and Phialemoniopsis pluriloculosa. The taxonomy of Phialemoniopsis has not been completely resolved, however, remaining incertae sedis within the Sordariomycetes. In addition, the new species Lecythophora luteorubra, Lecythophora cateniformis and Phialemonium globosum are described and the species Acremonium atrogriseum and Taifanglania inflata are transferred to the genus Phialemonium. Lecythophora and Phialemonium are currently monophyletic genera of the families Coniochaetaceae (Coniochaetales) and Cephalothecaceae (Sordariales) respectively, according to our results. Tables summarizing key morphological features to distinguish the current species of Lecythophora, Phialemonium and Phialemoniopsis are provided.

Cutaneous mucormycosis in the immunocompromised host: An important cause of persistent post traumatic skin lesions.

We describe a case of a 31-year-old man with a history of ocular non-Hodgkin's lymphoma who presented with a large 12-cm non-resolving traumatic skin lesion on his back. Biopsy showed fungal elements, and on fungal culture, Rhizopus arrhizus (formerly R. oryzae) was isolated. Cutaneous mucormycosis is an important diagnostic consideration for a non-resolving skin lesion in an immunocompromised host. Early tissue sampling is key, and diagnostic certainty is particularly important because first line therapy, liposomal amphotericin B, has significant systemic toxicities, notable renal toxicity, and is therefore challenging to continue empirically. Surgical debridement is an integral part of therapy, highlighting the need for early multidisciplinary care in patients with cutaneous mucormycosis.

The emergence of Trichophyton indotineae: Implications for clinical practice.

Emergence and increase of terbinafine-resistant dermatophytosis led to the identification of Trichophyton mentagrophytes internal transcriber space (ITS) genotype VIII in 2017, later renamed as Trichophyton indotineae and classified as a separate species in 2020. With its suspected origin in South Asia, this novel strain has emerged in Bahrain, Canada, Denmark, Finland, France, Germany, India, Iran, Japan, Russia, and Switzerland, with its spread attributed primarily to travel and migration. Diagnosis using routine mycology laboratory techniques is unable to distinguish T. indotineae from T. mentagrophytes and T. interdigitale; specific identification requires genomic sequencing to identify unique, specific markers. One speculated reason for this recent outbreak is the unrestricted use of topical steroid creams and antifungal agents. Patients with extensive tinea corporis and cruris due to T. indotineae present with inflammatory red plaques in multiple body sites. The majority of these infections prove to be resistant to conventional antifungals, including allylamines and azoles (itraconazole and fluconazole), thus emphasizing the need for antifungal susceptibility testing before treatment initiation and for reassessing in nonresponsive patients. Molecular studies have identified several point mutations in the ERG1 (terbinafine resistance) and ERG11 (azole resistance) genes, which need to be analyzed further. Use of relatively new agents, such as voriconazole and luliconazole, as well as device modalities and combination therapy, could be investigated for recalcitrant T. indotineae infections.

Monkey finger mycology? First case of otomycosis externa caused by Trichophyton simii after encounter with a monkey.

Unilateral ear pain, ear canal blockage and reduced hearing in an 18-year-old Canadian male who had travelled to India revealed, on examination of a swab, secretions bearing unusual fungal filaments visually suggestive of dermatophyte elements. Culture yielded Trichophyton simii, an unusual skin infecting species with a worldwide distribution but most often seen from India. The patient recalled swimming in the Ganges River but also had had his ear manipulated by a street monkey.

Utility of polymerase chain reaction for assessment of onychomycosis during topical therapy.

BACKGROUND: Increased detection of fungi including non-dermatophyte molds (NDMs) using polymerase chain reaction (PCR) methods is well-established. However, the use of PCR to evaluate ongoing onychomycosis treatment outcome has not been investigated. METHODS: Nail samples from 28 patients receiving topical efinaconazole were evaluated by both KOH/culture and PCR methods across the study period. Detection of microorganisms by PCR was compared to the culture at baseline and end of study at month 24 (M24). Fungal detection by both methods was evaluated with respect to clinical cure observed as 100% visual clearance of the target toenail. RESULTS: By culture, all 28 subjects were dermatophyte-positive at pre-treatment; only 4/28 also exhibited an NDM microorganism. According to PCR, 24/28 subjects were dermatophyte-positive pre-treatment, with 25/28 also exhibiting NDMs. At M24, 18/28 (64.3%) participants had negative KOH/culture results, in contrast to 4/28 (14.3%) negative PCR results. PCR showed higher rates of NDM detection than the culture at baseline as well as M24. Calculations to compare the diagnostic utility of KOH/culture versus PCR found that positive tests with both methods reliably indicate the presence of onychomycosis, but negative PCR correlated better with onychomycosis cure than did KOH/culture. DISCUSSION: PCR confirmed a high presence of NDMs pre-treatment, and continued presence of NDMs to M24, with unknown significance requiring further investigation. Though both KOH/culture and PCR have diagnostic limitations, PCR showed better overall utility than culture in predicting onychomycosis topical treatment outcome and should be more strongly considered for evaluation of topical therapies.

A Paradigm Shift in the Treatment and Management of Onychomycosis.

There is an increase in the incidence of onychomycosis, especially in at-risk populations. Onychomycosis is difficult to treat, as the efficacy of most antifungal agents is relatively low. Nondermatophyte molds (NDMs) and mixed infection (dermatophyte plus NDM) onychomycosis are contributing to growing antifungal resistance, as they are often underestimated and ignored due to incorrect diagnosis. There is a need for a paradigm shift in the management of onychomycosis to a patient-centered, holistic approach with an emphasis on laboratory diagnosis prior to initiating treatment, which enables the rational choice of the antifungal agent. Additionally, in the case of resistant infections, antifungal susceptibility testing is recommended. Strategies for effective management of onychomycosis include disinfection of fungal reservoirs in shoes and socks and prophylaxis posttreatment using topical antifungal agents. These measures may reduce the recurrence of onychomycosis and improve long-term clinical success.

The increasing problem of treatment-resistant fungal infections: a call for antifungal stewardship programs.

Antimicrobial stewardship (AMS) programs have been widely recognized among the public health community. These programs focus majorly on bacterial infections, efficient antibiotic use, and measures to curb increasing antibacterial resistance. AMS programs are successfully established around the globe; however, very few include antifungal stewardship (AFS). The increasing incidence of superficial and invasive fungal infections, combined with delayed or inaccurate diagnosis, has contributed to the overprescribing and overuse of antifungal agents. Such increased exposure to antifungal agents may be a reason for the emergence of increasing antifungal resistance among fungal pathogens. With mounting reports of treatment failures and resistant infections, the evidence to support the need for AFS programs is increasing. AFS is an emerging branch of AMS programs that requires global attention and recognition.

Internet-accessible DNA sequence database for identifying fusaria from human and animal infections.

Because less than one-third of clinically relevant fusaria can be accurately identified to species level using phenotypic data (i.e., morphological species recognition), we constructed a three-locus DNA sequence database to facilitate molecular identification of the 69 Fusarium species associated with human or animal mycoses encountered in clinical microbiology laboratories. The database comprises partial sequences from three nuclear genes: translation elongation factor 1α (EF-1α), the largest subunit of RNA polymerase (RPB1), and the second largest subunit of RNA polymerase (RPB2). These three gene fragments can be amplified by PCR and sequenced using primers that are conserved across the phylogenetic breadth of Fusarium. Phylogenetic analyses of the combined data set reveal that, with the exception of two monotypic lineages, all clinically relevant fusaria are nested in one of eight variously sized and strongly supported species complexes. The monophyletic lineages have been named informally to facilitate communication of an isolate's clade membership and genetic diversity. To identify isolates to the species included within the database, partial DNA sequence data from one or more of the three genes can be used as a BLAST query against the database which is Web accessible at FUSARIUM-ID (http://isolate.fusariumdb.org) and the Centraalbureau voor Schimmelcultures (CBS-KNAW) Fungal Biodiversity Center (http://www.cbs.knaw.nl/fusarium). Alternatively, isolates can be identified via phylogenetic analysis by adding sequences of unknowns to the DNA sequence alignment, which can be downloaded from the two aforementioned websites. The utility of this database should increase significantly as members of the clinical microbiology community deposit in internationally accessible culture collections (e.g., CBS-KNAW or the Fusarium Research Center) cultures of novel mycosis-associated fusaria, along with associated, corrected sequence chromatograms and data, so that the sequence results can be verified and isolates are made available for future study.

A dark strain in the Fusarium solani species complex isolated from primary subcutaneous sporotrichioid lesions associated with traumatic inoculation via a rose bush thorn.

Fusarium species are hyaline hyphomycetes widely distributed in nature and documented agents of both superficial and systemic infections in humans. In this paper, we report a darkly-pigmented and initially non-sporulating isolate in the Fusarium solani species complex (FSSC) causing a post-traumatic sporotrichoid infection in an otherwise healthy, male patient. Sequencing of multiple loci showed that the isolate represented an otherwise unknown lineage, possibly corresponding to a separate species, within the multi-species F. solani complex. In prolonged culture, the non-sporulating isolate produced revertant wild-type subcultures with typical Fusarium conidiation. This suggests that the original dense, dark, non-sporulating isolate was a host-adapted form selected in vivo for characters compatible with human pathogenicity. The production of such forms by Fusarium species is increasingly recognized now that sequencing has allowed the identification of highly atypical isolates. In vitro antifungal susceptibility of the isolate was investigated against seven conventional and two newly approved antifungal agents. The isolate showed in vitro resistance to amphotericin B, but appeared susceptible to itraconazole and terbinafine. A cure was ultimately achieved with combined terbinafine/itraconazole therapy with prolonged itraconazole follow-up therapy.

Phialosimplex, a new anamorphic genus associated with infections in dogs and having phylogenetic affinity to the Trichocomaceae.

Anamorphic members of the ascomycete family Trichocomaceae including Aspergillus, Penicillium, Paecilomyces, Geosmithia and Sagenomella have been reported from infections in canines. Six clinical isolates (five associated with infections in canines and one from a human source) demonstrated simple phialides producing conidia in long chains and were investigated for their potential relationship to Sagenomella chlamydospora, a known agent of canine disseminated mycosis. Phylogenetic analyses of internal transcribed spacer (ITS) and small subunit (SSU) region sequences revealed that all of the canine-associated isolates were distinct from Sagenomella species. The new anamorphic genus and species Phialosimplex caninus is described to accommodate the clinical isolates. Sagenomella chlamydospora and Sagenomella sclerotialis are transferred to the new genus as Phialosimplex chlamydosporus comb. nov. and Phialosimplex sclerotialis comb. nov.

The type isolate of a rare Phaeoacremonium species as a cause of human eumycetoma.

An isolate of unknown identity that had been identified as causing eumycetoma was retained in an international culture collection, and eventually became the nomenclatural type isolate of the rarely encountered Phaeoacremonium sphinctrophorum. The case featured an indurated, painless, swollen lesion on the dorsum of the foot that had developed in a Canadian resident who had previously been a farmer in Laos. Resection alone was curative.

High prevalence of mixed infections in global onychomycosis.

Onychomycosis is estimated at a prevalence of 10% worldwide with the infecting organism most commonly Trichophyton rubrum (T. rubrum). Traditional culture identification of causative organisms has inherent risks of overestimating dermatophytes, like T. rubrum, by inhibiting the growth of possible nondermatophyte mould (NDM) environmental contaminants which could be causative agents. Recently, molecular methods have revealed that a proportion of onychomycosis cases in North America may be caused by mixed infections of T. rubrum as an agent co-infecting with one or more NDM. Determining the global burden of mixed infections is a necessary step to evaluating the best therapies for this difficult-to-treat disease. To determine the prevalence of mixed infections in a global population, nail samples from onychomycosis patients in Brazil, Canada, and Israel (n = 216) were analyzed by molecular methods for the presence of dermatophytes and five NDMs. If an NDM was detected, repeat sampling was performed to confirm the NDM. T. rubrum was detected in 98% (211/216) of infections with 39% mixed (84/216). The infection type was more likely to be mixed in samples from Brazil, but more likely to be a dermatophyte in samples from Canada and Israel (Χ2 = 16.92, df = 2, P<0.001). The most common cause of onychomycosis was T. rubrum. In all countries (Brazil, Canada and Israel combined) the prevalence of dermatophyte (Χ2 = 211.15, df = 3, P<0.001) and mixed (dermatophyte and NDM; Χ2 = 166.38, df = 3, P<0.001) infection increased with patient age. Our data suggest that mixed infection onychomycosis is more prevalent than previously reported with the aging population being at increased risk for mixed infections.

Taxonomy and phylogeny of the Fusarium dimerum species group.

The morphospecies Fusarium dimerum, known only from its anamorph, comprises at least 12 phylogenetically distinct species. Analyses of the large subunit ribosomal DNA (LSU rDNA) show they are taxa of the Nectriaceae (Hypocreales), related to F. domesticum and form a phylogenetically distinct clade within Fusarium. Fusarium dimerum, for which no herbarium material could be located, is characterized by macroconidia with a single, median septum, according to the original description and illustration. Fusarium lunatum (= F. dimerum var. violaceum) forms similar but longer macroconidia and purple, catenate or clustered chlamydospores. Fusarium delphinoides sp. nov., F. biseptatum sp. nov., F. penzigii sp. nov., F. nectrioides comb. nov. (= F. dimerum var. nectrioides) and two unnamed Fusarium spp. produce macroconidia with mostly two or rarely three septa. The name F. dimerum, which originally was applied to a fungus from a citron, is used for a taxon including isolates causing infections in immunocompetent and immunocompromised patients. Fusarium nectrioides, F. delphinoides, F. penzigii and F. biseptatum are known from soil and dead plant substrata or rarely as agents of trauma-related eye infections of humans. Fusarium lunatum is an inhabitant of the cladodes of species within the cactus genera Opuntia and Gymnocalycium. Its unnamed closest sister taxon, which also forms 1-septate macroconidia and purple, clustered chlamydospores, was isolated from a human sinus. Fusarium delphinoides is a pathogen of the cactus-like African species Hoodia gordonii (Apocynaceae). Phylogenetic analyses based on combined sequences of the internal transcribed spacer region, LSU rDNA and partial sequences of the elongation factor 1-alpha and beta-tubulin genes identified a clade of several species producing predominately 2-septate macroconidia as the reciprocally monophyletic sister of F. dimerum. The basal sister group of the two aforementioned clades includes Fusarium lunatum and two undescribed species, all of which form 1-septate macroconidia.

Molecular mechanisms of pathogenicity: how do pathogenic microorganisms develop cross-kingdom host jumps?

It is common knowledge that pathogenic viruses can change hosts, with avian influenza, the HIV, and the causal agent of variant Creutzfeldt-Jacob encephalitis as well-known examples. Less well known, however, is that host jumps also occur with more complex pathogenic microorganisms such as bacteria and fungi. In extreme cases, these host jumps even cross kingdom of life barriers. A number of requirements need to be met to enable a microorganism to cross such kingdom barriers. Potential cross-kingdom pathogenic microorganisms must be able to come into close and frequent contact with potential hosts, and must be able to overcome or evade host defences. Reproduction on, in, or near the new host will ensure the transmission or release of successful genotypes. An unexpectedly high number of cross-kingdom host shifts of bacterial and fungal pathogens are described in the literature. Interestingly, the molecular mechanisms underlying these shifts show commonalities. The evolution of pathogenicity towards novel hosts may be based on traits that were originally developed to ensure survival in the microorganism's original habitat, including former hosts.